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|Application / Reactivity||Human|
|ELISA||6 ELISA Kits|
|Antigen||Wingless-Type MMTV Integration Site Family, Member 3A (WNT3A) ELISA Kits|
Kits with alternative reactivity to:
|Methode Type||Sandwich ELISA|
|Detection Range||0.156-10 ng/mL|
|Minimum Detection Limit||0.156 ng/mL|
|6 references available|
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Product Details WNT3A ELISA KitTarget details Application Details Handling References for WNT3A Kit (ABIN572397) Images
|Purpose||The kit is a sandwich enzyme immunoassay for in vitro quantitative measurement of WNT3A in Serum,Plasma,Tissue Homogenate,Cell Lysate,Biological Fluids|
|Sample Type||Serum, Plasma, Tissue Homogenate, Cell Lysate, Biological Fluids|
|Specificity||This assay has high sensitivity and excellent specificity for detection of Wingless Type MMTV Integration Site Family, Member 3A (WNT3A).|
|Cross-Reactivity (Details)||No significant cross-reactivity or interference between Wingless Type MMTV Integration Site Family, Member 3A (WNT3A) and analogues was observed.|
The sensitivity of this assay, or Lower Limit of Detection (LLD) was defined as the lowest protein concentration that could be differentiated from zero. It was determined by adding two standard deviations - the mean optical density value of twenty zero standard replicates and calculating the corresponding concentration.
|Characteristics||The test principle applied in this kit is Sandwich enzyme immunoassay. The microtiter plate provided in this kit has been pre-coated with an antibody specific to Wingless Type MMTV Integration Site Family, Member 3A (WNT3A). Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody specific to Wingless Type MMTV Integration Site Family, Member 3A (WNT3A). Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After TMB substrate solution is added, only those wells that contain Wingless Type MMTV Integration Site Family, Member 3A (WNT3A), biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450nm ± 10nm. The concentration of Wingless Type MMTV Integration Site Family, Member 3A (WNT3A) in the samples is then determined by comparing the O.D. of the samples to the standard curve.|
Pre-coated, ready to use 96-well strip plate: 1
Plate sealer for 96 wells: 4
Standard Diluent: 1×20 mL
Detection Reagent A: 1×120 µL
Assay Diluent: A 1×12 mL
Detection Reagent B: 1×120 µL
Assay Diluent B: 1×12 mL
Substrate A: 1×10 mL
Substrate B: 1×2 mL
Wash Buffer (30 × concentrate): 1×20 mL
Instruction manual: 1
|Material not included||
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|Alternative Name||WNT3A (WNT3A ELISA Kit Abstract)|
Application DetailsProduct Details WNT3A ELISA Kit Target details Handling References for WNT3A Kit (ABIN572397) Images back to top
Information on standard material:
|Sample Volume||100 μL|
|Assay Time||4.5 h|
|Plate||Pre-coated,Strips (12 x 8),96 wells|
1. Prepare all reagents, samples and standards
2. Add 100 µL standard or sample to each well. Incubate 2 hours at 37 °C
3. Aspirate and add 100 µL prepared Detection Reagent A. Incubate 1 hour at 37 °C
4. Aspirate and wash 3 times
5. Add 100 µL prepared Detection Reagent B. Incubate 30 minutes at 37 °C
6. Aspirate and wash 5 times
7. Add 90 µL Substrate Solution. Incubate 15-25 minutes at 37 °C
8. Add 50 µL Stop Solution. Read at 450 nm immediately.
Serum: Allow samples to clot for two hours at room temperature or overnight at 4°C before centrifugation for 20 minutes at approximately 1000 × g. Assay immediately or store samples in aliquot at -20°C or -80°C. Avoid repeated freeze/thaw cycles.
Plasma: Collect plasma using EDTA or heparin as an anticoagulant. Centrifuge samples for 15 minutes at 1000 × g within 30 minutes of collection. Remove plasma and assay immediately or store samples in aliquot at -20°C or -80°C. Avoid repeated freeze/thaw cycles.
Tissue Homogenates: The preparation of tissue homogenates will vary depending upon tissue type. For this assay, rinse tissues in ice-cold PBS (0.02mol/L,pH 7.0-7.2) to remove excess blood thoroughly and weigh before homogenization. Mince the tissues to small pieces and homogenize them in 5-10 mL of PBS with a glass homogenizer on ice (Micro Tissue Grinders work, too). Sonicate the resulting suspension with an ultrasonic cell disrupter or subject it to two freeze-thaw cycles to further break the cell membranes. Centrifugate the homogenates for 5 minutes at 5000 × g. Remove the supernate and assay immediately or aliquot and store at -20°C
Cell Lysate: Cells must be lysed before assaying according to the following directions. Adherent cells should be detached with trypsin and then collected by centrifugation (suspension cells can be collected by centrifugation directly). Wash cells three times in cold PBS. Resuspend cells in PBS (1×) and subject them to ultrasonication for 4 times (or Freeze cells at -20 °C. Thaw cells with gentle mixing. Repeat the freeze/thaw cycle for 3 times.) Centrifuge at 1500 × g for 10 minutes at 2 - 8°C to remove cellular debris.
Biological Fluids: Centrifuge samples for 20 minutes at 1000 × g. Remove particulates and assay immediately or store samples in aliquot at -20 °C or -80 °C for later use. Avoid repeated freeze/thaw cycles.
|Calculation of Results||
Average the duplicate readings for each standard, control, and samples and subtract the average zero standard optical density. Construct a standard curve by plotting the mean O.D. and concentration for each standard and draw a best fit curve through the points on the graph or create a standard curve on log-log graph paper with WNT3A concentration on the y-axis and absorbance on the x-axis. Using some plot software, for instance, curve expert 1.30, is also recommended. If samples have been diluted, the concentration read from the standard curve must be multiplied by the dilution factor.
In order to make the calculation easier, we plot the O.D. value of the standard (X-axis) against the known concentration of the standard (Y-axis), although concentration is the independent variable and O.D. value is the dependent variable. However, the O.D. values of the standard curve may vary according to the conditions of assay performance (e.g. operator, pipetting technique, washing technique or temperature effects), plotting log of the data to establish standard curve for each test is recommended. Typical standard curve below is provided for reference only.
Intra-assay Precision (precision within an assay): 3 samples with low, middle and high level human WNT3A were tested 20 times on one plate, respectively.
Inter-assay Precision (precision between assays): 3 samples with low, middle and high level human WNT3A were tested on 3 different plates, 8 replicates in each plate.
CV (%) = SD/mean X 100
|Restrictions||For Research Use only|
HandlingProduct Details WNT3A ELISA Kit Target details Application Details References for WNT3A Kit (ABIN572397) Images back to top
|Precaution of Use||The Stop Solution suggested for use with this kit is an acid solution. Wear eye, hand, face, and clothing protection when using this material.|
|Handling Advice||The stability of kit is determined by the loss rate of activity. The loss rate of this kit is less than 5 % within the expiration date under appropriate storage condition. To minimize extra influence on the performance, operation procedures and lab conditions, especially room temperature, air humidity, incubator temperature should be strictly controlled. It is also strongly suggested that the whole assay is performed by the same operator from the beginning to the end.|
|Storage||4 °C/-20 °C|
|Expiry Date||6 months|
References for WNT3A Kit (ABIN572397)Product Details WNT3A ELISA Kit Target details Application Details Handling Images back to top
|Product cited in:||
Furuya, Sasaki, Tsunoda, Tsuji, Udaka, Oyamada, Tsuchiya, Oguchi: "Eribulin upregulates miR-195 expression and downregulates Wnt3a expression in non-basal-like type of triple-negative breast cancer cell MDA-MB-231." in: Human cell, Vol. 29, Issue 2, pp. 76-82, 2016
Pan, Yao, Zheng, Gu, Yang, Qiu, Cai, Wu, Yao: "Abnormality of Wnt3a expression as novel specific biomarker for diagnosis and differentiation of hepatocellular carcinoma." in: Tumour biology, Vol. 37, Issue 4, pp. 5561-8, 2016
Lu, Li, Liu, Wu, Zhang, Xie, Wang: "Wls promotes the proliferation of breast cancer cells via Wnt signaling." in: Medical oncology (Northwood, London, England), Vol. 32, Issue 5, pp. 140, 2015
Lin, Ueng, Niu, Yuan, Yang, Chen, Lee, Chen: "Effects of hyperbaric oxygen on the osteogenic differentiation of mesenchymal stem cells." in: BMC musculoskeletal disorders, Vol. 15, pp. 56, 2014
Klingberg, Nurkkala, Carlsten, Forsblad-dElia: "Biomarkers of bone metabolism in ankylosing spondylitis in relation to osteoproliferation and osteoporosis." in: The Journal of rheumatology, Vol. 41, Issue 7, pp. 1349-56, 2014
Nazarian, Ghaffari Novin, Jalili, Mirfakhraie, Heidari, Hosseini, Norouzian, Ehsani: "Expression of Glycogen synthase kinase 3-? (GSK3-?) gene in azoospermic men." in: Iranian journal of reproductive medicine, Vol. 12, Issue 5, pp. 313-20, 2014
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